Phospholipids with a glycerol-phosphorylglycerol backbone are associated with lysosomes and are present at elevated amounts in mammalian cells containing excessive numbers of secondary lysosomes. In the cellular slime mold, Dictyostelium discoideum, such phospholipids are believed to be associated with digestive food vacuoles, which are secondary lysosomes used to digest bacteria taken up by phagocytosis. Preliminary studies on the structure of the phospholipids indicate that they are bisphosphatidic acid (BPA), semi-lysobisphosphatidic acid, and their respective plasmalogen forms. In this investigation, the preliminary studies will be extended and the structures of the phospholipids will be determined by a combination of chemical and spectral techniques. Since both BPA and food vacuoles accumulate in cells growing on bacteria and disappear in aggregating cells, the relationship of BPA biosynthesis to the formation of food vacuoles and the relationship of BPA catabolism to food vacuoles disappearance will be examined. The regulation of BPA metabolism will be investigated by subjecting cells to conditions in which BPA accumulates and disappears. The mechanism of BPA synthesis and catabolism will be studied using isotope tracer methods. Using BPA as a marker for food vacuoles, the proposed studies will provide new insights on 1. the formation of secondary lysosomes produced as lysosomes fuse with phagocytic vesicles derived from the plasma membrane; and 2. the degradation of secondary lysosomes as they fuse with the plasma membrane and release their contents by exocytosis.